1. Field of the Invention
The present invention relates to a new gene, a part of which encodes a polypeptide having a lymphotoxin activity, a method for the production of the gene, and a human lymphotoxin produced from the gene by using genetic engineering technology.
The present invention also relates to an antitumor composition comprising the lymphotoxin as an active ingredient.
2. Description of the Background Art
Lymphotoxin (hereinafter referred to as LT) is known to be a kind of lymphokine which is specifically or non-specifically released from lymphocytes or established lymphoid cell lines and which has a cytotoxic activity. LT not only has a cytotoxic activity against various cancer cells but also an activity for intensifying the cytotoxic activity of certain kinds of carcinostatics or interferons, and it is expected to be usable as a tumoricidal agent or as other types of medicinal agents. (Refer to Granger G. A. et al., "International Congress of Chemotherapy", Kyoto, Japan, June 23-28, Abstracts, p. 15; and Matsunage K. et al., ibid, p. 352.)
For the production of a human LT, a variety of methods have been known where human-derived cells or human-derived tumor cells are cultured and the cultured supernatant liquid is purified to obtain the human LT. For instance, a method is known wherein tonsil cells or peripheral blood lymphocytes are cultured together with a phytohemagglutinin (hereafter referred to as PHA) and the human LT is isolated from the cultured supernatant liquid (refer to Peter, T. B. et al., J. Immunol., 111, 770 (1973); Walker, S. M. and Lucas, Z. J., J. Immunol., 109, 1233 (1972)). Another method is known wherein lymphoid tumor cells are cultured and the human LT is isolated from the cultured supernatant liquid (refer to Yamamoto, R. S. et al., J. Biol. Response Modifiers, 3, 76 (1984); European Patent Application No. 0100641). Yet another method is known wherein T-cell hybridomas are cultured in the presence of phorbol myristate acetate (hereinafter referred to as PMA) and/or concanavalin A (hereinafter referred to as Con A) and the human LT is isolated from the cultured supernatant liquid (refer to Asada, M. et al., Cell. Immunol., 77, 150 (1983)).
These known methods, however, have problems in that the content of LT contained in the culture supernatant solution is extremely small and the nutrient sources to be used in the cultivation (for example, fetal calf serum) are expensive. Therefore, economical production of LT of high purity is difficult by the known methods.
For the production of human LT, it would be desirable to have a means employing genetic engineering technology, wherein a gene corresponding to LT is inserted into a vector and the resulting recombinant plasmid is replicated, transcribed and translated in bacteria, fungi, yeasts or animal cells in order to obtain the desired human LT as produced in these cells.
The present inventors have previously obtained an LT-producing human T-cell hybridoma clone A-C5-8 cell line by the use of a human T-cell hybridization method (emetine/actinomycin-D method). (Refer to Asada, M., et al., Cell Immunol., 77, 150 (1983).) However, when the A-C5-8 cell line was cultured in the presence of Con A and PMA under the optimum LT-producing condition (incubation period of 30 hours or more), the messenger RNA (hereinafter referred to as mRNA) corresponding to LT could not be obtained, and therefore, the gene corresponding to LT could not be obtained. Thus, a need continues to exist for a method of producing LT by way of genetic engineering technology.